The major goal of this proposal is to investigate the role of the polyphosphoinositide-protein-kinase-C (PI-PKC) second-messenger system in the mammalian brain using the hippocampus as a model system. A number of important functions have been suggested for this system, including regulation of calcium- and potassium- dependent currents, transmitter release and long-term potentiation. Until now the system has primarily been studied using phorbol esters, plant-derived tumor- promoting compounds that activate PKC. Of particular interest in this phase of the project is the role of endogenous activation of PKC by acetylcholine and its analogs. Little is known about physiological effects of neurotransmitter activation of the PI PKC system. Electrophysiological responses of hippocampal neurons to cholinergic agents will be studied with intracellular recording in the slice preparation. In acutely dissociated and tissue-cultured neurons, whole-cell voltage- and patch-clamp techniques will be used. Direct activation of PKC by synthetic diacylglycerols and phorbol esters, in addition to direct intracellular injection of purified PKC, will be used to define which ionic currents are affected and how they are affected. We will also study the responses to injection of the other product of polyphosphoinositide breakdown, inositol-1,4,5-triphosphate. A variety of the known and suspected consequences of PKC activation imply that it is involved in control of neuronal excitability under normal circumstances. Malfunction in excitability control is a major problem in several neurological diseases, including epilepsy, Parkinson's and Huntington's diseases. Deficits in cholinergic systems have been implicated in Alzheimer's disease. Defining the roles of the PI-PKC system in excitability control may not only lead to better understanding of the disease states, but also contribute to rational design of drug therapy.